Alzheimer's disease exhibits a symptom of progressive dementia. Its pathologic histology is characterized by emergence of a huge number of senile plaques in the brain and accumulation of neurofibrillary degenerations in neurons. The disease is neurodegenerative in which neurons are gradually leading to deciduation. Alzheimer's disease generally develops in old age and its prevalence is known to increase with aging. At present, a definitive treatment of Alzheimer's disease is impossible. Accordingly, in order to prepare for sharp increase of the old age population in the future, early developments of a method of therapeutic and preventive treatment of Alzheimer's disease and an effective medicament for preventive and therapeutic treatment of the disease are desired.
Senile plaque is a deposit outside neurons which contains various ingredients, and whose main ingredient is a peptide consisting of 39–42 amino acid residues called amyloid β protein (Aβ). Amyloid precursor protein (APP) is cleaved by proteases tentatively named β secretase and γ secretase to produce amyloid β. In the senile plaque, the amyloid β deposits as a rigid construct having β sheet structure. The senile plaque is first formed as a “stain-like” deposition called as a diffuse senile plaque. At this stage, neurodegeneration has not yet occurred. It is considered that, as the diffuse senile plaque becomes a more rigid deposition, the degeneration or deciduation of neurocytes occurs, which results in the onset of symptoms of Alzheimer's disease such as dementia. There are Aβ 40 consisting of 40 amino acid residues and Aβ 42 consisting of 42 amino acid residues as main amyloid β. Most of amyloid β generated by cells is Aβ 40, and only a little amount of Aβ 42 exists. However, Aβ 42 has higher aggregation properties, and therefore, A β42 is considered to have a more significant role than Aβ 40 in the formation of senile plaque (Tamaoka, Naika (Internal Medicine), Vol. 77, P843, 1996).
In Alzheimer's disease, familial onsets are observed which exhibit an autosomal dominant inheritance. A gene first identified as a causal gene of the familial onset of Alzheimer's disease in 1991 is a mutant of APP, a gene located on chromosome 21 in which amino acid residue at position 717 is mutated from valine to isoleucine (Goate A. et al., Nature, Vol. 349, P704, 1991).
Other mutants of APP as causes of Alzheimer's disease were found such as those where said amino acid residue at position 717 is mutated to phenylalanine (Murrell J. et al., Science, Vol. 254, P97, 1991); where the amino acid residue at the same position is mutated to glycin (Chartier, Harlin et al., Nature, Vol. 353, P844, 1991); where two amino acid residues at positions 670 and 671 are mutated from lysine-methionine to asparagine-leucine (Mullan M. et al., Nature Genet., Vol. 1, P345, 1992); and where amino acid residue at position 692 is mutated from alanine to glycin (Hendrisk L. et al., Nature Genet., Vol. 1, P218, 1992) and the like.
Apolipoprotein E (apo E) was reported in 1993 as a causal factor or a risk factor of the familial Alzheimer's disease. Persons with Alzheimer's disease were found to have apoE4, in which the amino acid residue at position 112 is arginine and the amino acid residue at position 158 is arginine, at a significantly higher rate than healthy persons among isomers of apoE whose genes are located on chromosome 19 (Corder E. H. et al., Science, Vole 261, P921, 1993).
After then, a mutant of the gene “presenilin-1” (PS-1, initially called as S182) being located on chromosome 14 (Sherrington R. et al., Nature, Vol. 375, P754, 1995) and a mutant of the gene “presenilin-2” (PS-2, initially called as E5-1 or STM-2) being located on chromosome 1 (Sherrington R. et al., Nature, Vol. 375, P754, 1995) were found as new causal genes for Alzheimer's disease in 1995 (in the specification, each gene is called as “presenilin-1 gene” and “presenilin-2 gene”, respectively, and each gene product is called as “presenilin-1 protein” and “presenilin-2 protein”, or “PS-1” and “PS-2”, respectively.)
Presenilin-1 protein and presenilin-2 protein consisting respectively of 467 and 448 amino acid residues have a seven (or eight)-fold transmembrane primary structure, and accordingly, they are presumably present as membrane proteins. Homology of the two proteins is high at amino acids level, i.e., 67% in total and 84% in the transmembrane domain alone. As for function of presenilin-1 protein, the protein is suggested to possibly have similar functions to nematode sel-12 protein or SPE-4 protein because of high homology to these proteins. SPE-4 protein participates in nematode spermatogenesis process and is considered to be involved in transport and storage of proteins.
Consequently, presenilin-1 protein is believed to participate possibly in processing of membrane proteins such as APP, axoplasmic transport, and fusion of membrane vesicle with membranes. The sel-12 was found as a gene which remedies an embryological abnormality caused by mutation of lin-12 which controls nematode development. The lin-12 is considered to be involved in intercellular signal transduction, and accordingly, presenilin-1 protein is also suggested to possibly participate in a certain step of intercellular signal transduction.
The first report on presenilin-1 protein describes that mutations causing the familial Alzheimer's disease are substitutions of amino acid residues at five positions. After this report, genes mutated at various sites were found from many families afflicted with familial Alzheimer's disease, which include OS-2 (isoleucine at position 213 is mutated to threonine) and OS-3 (valine at position 96 is mutated to phenylalanine), both reported by the present inventors (Kamino K. et al., Neurosci., Lett., Vol. 208, P195, 1996), and more than 40 types of amino acid substitutions have been known at more than 30 sites so far (Hardy. TINS, Vol. 20, P154, 1997).
At present, 70–80% of the familial Alzheimer's disease is believed to be related to the mutation of presenilin-1 protein. Mutations at two sites have been reported as for presenilin-2 protein. As explained above, genetic analysis has proved that mutants of presenilin-1 and presenilin-2 proteins are deeply involved in the familial Alzheimer's disease.
Studies on mechanism how the mutants of presenilin-1 and presenilin-2 proteins cause the onset of Alzheimer's disease have also been progressed. It has been reported that Aβ 40 is almost the same level as normal presenilin-1 and presenilin-2 proteins, whilst Aβ 42 is highly increased as compared to normal presenilin-1 and presenilin-2 proteins in serum or a culture medium of dermal fibroblasts from a patient with Alzheimer's disease having the aforementioned mutants (Scheuner D. et al.: Nature Med., Vol. 2, P864, 1996); in a culture medium of a cell line transformed by mutants of presenilin-1 protein and presenilin-2 protein (Xia W. et al.: J. Biol. Chem. Vol. 272, P7977, 1997; Borchelt D. R. et al.: Neuron, Vol. 17, P1005, 1996; Citron, M. et al.: Nature Med., Vol. 3, P67, 1997); and in the brain tissue of a patient with familial Alzheimer's disease having the mutant presenilin-1 protein (Lemere C. A. et al.: Nature Med., Vol. 2., P1146, 1996).
These reports show that the mutants of presenilin-1 protein and presenilin-2 protein, which cause the familial Alzheimer's disease, possibly trigger the onset of Alzheimer's disease by the increase of A β′42 which is considered to play a significant role in the formation of senile plaque. A trans-genic mouse transferred with a gene encoding the mutant presenilin-1 protein was created (Duff K. et al.: Nature, Vol. 383, P710, 1996, Borchelt D R. et al.: Neuron, Vol. 17, P1005, 1996 and Citron M. et al.: Nature Med., Vol. 3, P67, 1997). It was reported that Aβ 42 in the brain of the trans-genic mouse selectively increased. These results are strong supports of the possibility that mutants of presenilin-1 protein and presenilin-2 protein causing the familial Alzheimer's disease increase Aβ 42 which possibly has significant roles in the formation of senile plaque, thereby develop Alzheimer's disease. However, no description is given about histological study of the mouse's brain in the above reports on the trans-genic mouse, which presumably due to no observation of remarkable histological change in the brain of the trans-genic mouse.
Generally, trans-genic animals are useful as a means of analyzing functions of a target gene in vivo. However, it is technically difficult to control the expression of a transferred gene quantitatively, tissue specifically, or time specifically during development. There is also a problem in that two different gene products are present as a mixture in the trans-genic animals since a gene inherently possessed by the animal still works for normal expression, and functions of a transferred gene cannot be sufficiently analyzed. Furthermore, when the transferred gene is subjected to particularly excessive expression, functions not inherently performed in vivo may appear in trans-genic animals, which results in a defect of possible confusion in analysis of constructed gene-mutated animals.
Apart from trans-genic animals, knockout animals may also be used as a means of analyzing functions of a target gene. In a knockout animal, a target gene inherently possessed by the animal is artificially destroyed so as to be dysfunctional. A detailed analysis of knockout animals may reveal functions of a target gene in vivo. However, particular changes in knockout animals created as homozygote sometimes fails to appear, since the functions of the other gene products in the knockout animal may substitute for that of the destroyed gene products. Furthermore, there is also a problem in that an animal as homozygote may sometimes be lethal because the destroyed gene product is essential to the animal's development and growth, whilst thorough analysis of gene functions of an animal as viable heterozygote is practically impossible.